The purposes of this project are 1) to clarify the factors that affect hyaluronan-depolymerizing enzyme which degrade high-molecular-weight hyaluronan into fragments with molecular weight of 40,000, and 2) to characterize the hyaluronan-deficient extracellular matrix which were formed by cultivating tcells in the presence of 4-methylimbelliferone, an inhibitor of hyaluronan synthase. First, heparitinase treatment of the cell layer of cultured fibroblasts resulted the stimulation of the depolymerization of exogenous hyaluronan. Heparitinase treatment also showed the inhibition of the synthesis of high-molecular-weight hyaluronan. These results suggest that the cell surface heparan sulfate may affect hyaluronan metabolisim. Second, the analysis of hyaluronan-deficient extracellular matrix, which can be formed by cultivation of the cells in the presence of 0.5 mM 4-methylumbelliferone, showed the decrease of CD44 and fibronectin in cell layer. The amount of type I collagen in cell layer was not changed, while an increase of the molecule was observed in the medium by 4-methylumbelliferone. These phenomena may be caused by the lack of hyaluronan in extracellular matrix. Furthermore, the relation between hyaluronan and matrix metalloproteinases were investigated. Cells were cultured in the presence of exogenous hyaluronan and the activities of matrix metalloproteinases were compared with that of control cells by zymography and western blotting. As a result, the excretion of stromelysin-1 was found to be strongly suppressed by high-molecular weight hyaluronan with high-concentration of more than 1 mg/ml.